Presently, two Gi/o, protein coupled cannabinoid receptors have been characterized in mammals and other organisms (Devane et al, Brain Mol. Pharmacol. (1988) 34: 605-613, Matsuda et at, Nature (1990) 346: 561-564). Cannabinoid receptor 1 (CB1) is a central receptor found in the mammalian brain and a number of other sites in peripheral tissues; and CB2, a peripheral receptor found principally in cells related to the immune system. Cannabinergic ligands can bind to the CB1 and/or CB2 receptors in an individual or animal. In vitro methods for assaying the ability of a compound to bind to CB1 and/or CB2 receptors are known (Devane et al, Brain Mol. Pharmacol. (1988) 34: 605-613). Results from the in vitro assay correlate with and predict the in vivo ability of that compound to bind to CB1 and/or CB2 receptors and modulate their function(s).
When introduced in an individual or animal some of these cannabinergic ligands can bind to and directly modulate (activate or deactivate) the CB1 and/or CB2 receptors. Many physiological effects have been associated with direct modulation of the CB1 and/or CB2 receptors in an individual or animal (Jonsson et al. Basic and Clinical Pharmacol. Toxic. (2006) 98: 124-134), Examples of cannabinergic ligands include (−)-Δ9-tetrahydrocannabinol ((−)-Δ9-THC), the principal bioactive constituent of cannabis and exogenous ligand for the cannabinoid CB1 and CB2 receptors) and other synthetic cannabinergic analogs. The major endogenous ligands for the CB receptors are N-arachidonoyl ethanolamine (anandamide, AEA) and 2-arachidonoylglycerol (2-AG) (Devane et al, Science (1992) 258: 1946-1949, Mechoulam et al Biochem. Pharmacol. (1995) 50: 83-90).
The magnitude and duration of in vivo CB1 and/or CB2 receptor modulation by endocannabinoids AEA and 2-AG is relatively short, presumably due to rapid inactivation process involving endocannabinoid deactivating proteins. Anandamide for example, is inactivated via fatty acid amide hydrolase (FAAH) mediated hydrolysis (Deutsch et al. Biochem. Pharmacol. 1993, 46, 791-796). FAAH belongs to the amidase signature (AS) super family of serine hydrolases and in contrast to the classical serine-histidine-aspartate triad found in most serine hydrolases, the catalytic machinery of this enzyme is a serine-serine-lysine catalytic triad. FAAH has been isolated, molecularly cloned and its 2.8 Å crystal structure was recently reported. 2-arachidonoylglycerol (2-AG), 1-arachidonoylglycerol, arachidonamide and the corresponding simple ester methyl arachidonate are also substrates for FAAH.
Moreover, studies have demonstrated that this enzyme not only can hydrolyze anandamide into arachidonic acid and ethanolamine it can also catalyze reverse synthesis from the two hydrolysis components. Also notable is FAAH's ability to hydrolyze several bioactive fatty acid amides not belonging to the endocannabinoid family, for example, the sleep inducing lipid oleamide, the appetite-suppressing agent oleoylethanolamine, the related. 1-oleoylglycerol, and the peripheral analgesic and anti-inflammatory mediator palmitoylethanolamine. Despite the fact that a range of fatty acid amides, ethanolamides and esters are hydrolyzed by FAAH, this enzyme appears to work most effectively on arachidonoyl and oleoyl substrates.
Although FAAH has been shown to also catalyze hydrolysis of 2-arachidonoylglycerol in vitro, a distinct enzyme, monoacylglycerol lipase (MAGL) plays the predominant role in catalyzing 2-AG (the most abundant endocannabinoid) hydrolysis in vivo (Karlsson et al J. Biol. Chem. (1997) 272: 27218-27223). Monoacylglycerol lipase, also known as MAGL, MAG lipase or MAGL is a serine hydrolase that is also notable for its ability to hydrolyze several bioactive fatty acid glyceryl esters not belonging to the endocannabinoid family, for example, 2-oleoylglycerol and 2-palmitoyl glycerol. MAGL plays dual roles in physiologic processes, encompassing regulating endocannabinoid tone as well as lipogenesis (Dinh et al Proceedings of the National Academy of Sciences of the United States of America (2002) 99: 10819; Schlosburg et al Nature neuroscience (2010) 13: 1113. Site-directed mutagenesis studies (Zvonok et at Chemistry & biology (2008) 15: 854; Zvonok et at Journal of Proteome Research (2008) 7: 2158) as well as X-ray crystal structure of a complex with its ligand have identified the enzyme's catalytic triad as Ser122-Asp239-His269 (Labar et at Chembiochem: a European journal of chemical biology (2010) 11: 218; Schalk-Hihi et at Protein science: a publication of the Protein Society (2011) 20: 670.
Some compounds can inhibit the inactivation of cannabinergic ligands by FAAH, by MAGL, or by dual FAAH/MAGL. These compounds may not bind to, or may have lesser affinity for, the cannabinoid receptors. Thus, the physiological action for such compounds is inhibition of fatty acid amide hydrolase (FAAH) and not direct modulation of the CB′ and/or CB2 receptors. The inactivation of endocannabinoids by FAAH, MAGL, or dual FAAH/MAGL can be inhibited. These inhibitors may not bind to, or may have lesser affinity for, the cannabinoid receptors. Thus, the physiological action for such compounds is inhibition of FAAH and or MAGL and not direct modulation of the CB1 and/or CB2 receptors. Inhibition of FAAH, MAGL, or dual FAAH/MAGL in an individual or animal will slow the normal degradation and inactivation of endogenous cannabinoid ligands by FAAH or MAGL hydrolysis and allow higher levels of those endogenous cannabinergic ligands to remain present in that individual or animal. These higher levels of endocannabinoid induce increased stimulation of the cannabinoid CB1 and CB2 receptors and produce physiological effects related to the activation of the cannabinoid receptors. They will also enhance the effects of other exogenous cannabinergic ligands and allow them to produce their effects at lower concentrations as compared to systems in which FAAH, MAGL, or dual FAAH/MAGL action is not inhibited. Thus, a compound that inhibits the inactivation of endogenous cannabinoid ligands by FAAH, MAGL, or dual FAAH/MAGL may increase the levels of endocannabinoids and, thus, enhance the activation of cannabinoid receptors. The compound may not directly modulate the cannabinoid receptors but has the effect of indirectly stimulating the cannabinoid receptors by increasing the in vivo levels of endocannabinoid ligands. It may also enhance the effects and duration of action of other exogenous cannabinergic ligands that are administered in order to elicit a cannabinergic response.
Marijuana-like cannabinoids, in addition to acting at cannabinoid receptors also affect cellular membranes, thereby producing undesirable side effects such as drowsiness, impairment of monoamide oxidase function and impairment of non-receptor mediated brain function. The addictive and psychotropic properties of some cannabinoids also limit their therapeutic value. Compounds that inhibit FAAH, MAGL, or dual FAAH/MAGL activity may indirectly provide desirable pharmacological properties while avoiding the disadvantages incurred by use of cannabinergic ligands that directly activate the cannabinoid receptors. Compounds that inhibit FAAH and or MAGL activity provide an alternative mechanism for indirectly stimulating cannabinoid receptors and may provide desirable pharmacological properties without the addictive and psychotropic properties as well as other undesirable properties associated with exogenous cannabinergic ligands.
FAAH, MAGL, or dual FAAH/MAGL inhibitory compounds comprise two pharmacophoric subunits responsible for enzyme recognition and inactivation. The “inhibition” subunit typically comprises an activated carbonyl group and the “binding” subunit, which is linked to the inhibition subunit, enhances the inhibitory action of the molecule.
Conditions that may be treated by modulation of the CB1/CB2 cannabinoid receptors include for example: high blood pressure disease or hypertension; peripheral vascular diseases; coronary artery disease; abnormal heart rate; pulmonary hypertension; ocular hypertension or glaucoma; tumor growth; to prevent or reduce inflammation; to provide neuroprotection; to treat epilepsy; to treat nausea, such as associated with cancer chemotherapy; AIDS wasting syndrome as well as other ailments in which cannabinoid system is implicated.